Post-Messinian evolutionary relationships across the Sicilian channel: Mitochondrial and nuclear markers link a new green toad from Sicily to African relatives

Background

The major birch pollen allergen, Bet v 1, is a member of the ubiquitous PR-10 family of plant pathogenesis-related proteins. In recent years, a number of diverse plant proteins with low sequence similarity to Bet v 1 was identified. In addition, determination of the Bet v 1 structure revealed the existence of a large superfamily of structurally related proteins. In this study, we aimed to identify and classify all Bet v 1-related structures from the Protein Data Bank and all Bet v 1-related sequences from the Uniprot database.

Results

Structural comparisons of representative members of already known protein families structurally related to Bet v 1 with all entries of the Protein Data Bank yielded 47 structures with non-identical sequences. They were classified into eleven families, five of which were newly identified and not included in the Structural Classification of Proteins database release 1.71. The taxonomic distribution of these families extracted from the Pfam protein family database showed that members of the polyketide cyclase family and the activator of Hsp90 ATPase homologue 1 family were distributed among all three superkingdoms, while members of some bacterial families were confined to a small number of species. Comparison of ligand binding activities of Bet v 1-like superfamily members revealed that their functions were related to binding and metabolism of large, hydrophobic compounds such as lipids, hormones, and antibiotics. Phylogenetic relationships within the Bet v 1 family, defined as the group of proteins with significant sequence similarity to Bet v 1, were determined by aligning 264 Bet v 1-related sequences. A distance-based phylogenetic tree yielded a classification into 11 subfamilies, nine exclusively containing plant sequences and two subfamilies of bacterial proteins. Plant sequences included the pathogenesis-related proteins 10, the major latex proteins/ripening-related proteins subfamily, and polyketide cyclase-like sequences.

Conclusion

The ubiquitous distribution of Bet v 1-related proteins among all superkingdoms suggests that a Bet v 1-like protein was already present in the last universal common ancestor. During evolution, this protein diversified into numerous families with low sequence similarity but with a common fold that succeeded as a versatile scaffold for binding of bulky ligands.     

Sequestration and biosynthesis of cyanogenic glucosides in passion vine butterflies and consequences for the diversification of their host plants

The colorful heliconiine butterflies are distasteful to predators due to their content of defense compounds called cyanogenic glucosides (CNglcs), which they biosynthesize from aliphatic amino acids. Heliconiine larvae feed exclusively on Passiflora plants where ~30 kinds of CNglcs have been reported. Among them, some CNglcs derived from cyclopentenyl glycine can be sequestered by some Heliconius species. In order to understand the evolution of biosynthesis and sequestration of CNglcs in these butterflies and its consequences for their arms race with Passiflora plants, we analyzed the CNglc distribution in selected heliconiine and Passiflora species. Sequestration of cyclopentenyl CNglcs is not an exclusive trait of Heliconius, since these compounds were present in other heliconiines such as Philaethria, Dryas and Agraulis, and in more distantly related genera Cethosia and Euptoieta. Thus, it is likely that the ability to sequester cyclopentenyl CNglcs arose in an ancestor of the Heliconiinae subfamily. Biosynthesis of aliphatic CNglcs is widespread in these butterflies, although some species from the sara‐sapho group seem to have lost this ability. The CNglc distribution within Passiflora suggests that they might have diversified their cyanogenic profile to escape heliconiine herbivory. This systematic analysis improves our understanding on the evolution of cyanogenesis in the heliconiine–Passiflora system.     

Reduced Rates of Sequence Evolution of Y-Linked Satellite DNA in Rumex (Polygonaceae)

One characteristic of sex chromosomes is the accumulation of a set of different types of repetitive DNA sequences in the Y chromosomes. However, little is known about how this occurs or about how the absence of recombination affects the subsequent evolutionary fate of the repetitive sequences in the Y chromosome. Here we compare the evolutionary pathways leading to the appearance of three different families of satellite-DNA sequences within the genomes of Rumex acetosa and R. papillaris, two dioecious plant species with a complex XX/XY₁Y₂ sex-chromosome system. We have found that two of these families, one autosomic (the RAE730 family) and one Y-linked (the RAYSI family), arose independently from the ancestral duplication of the same 120-bp repeat unit. Conversely, a comparative analysis of the three satellite-DNA families reveals no evolutionary relationships between these two and the third, RAE180, also located in the Y chromosomes. However, we have demonstrated that, regardless of the mechanisms that gave rise to these families, satellite-DNA sequences have different evolutionary fates according to their location in different types of chromosomes. Specifically, those in the Y chromosomes have evolved at half the rate of those in the autosomes, our results supporting the hypothesis that satellite DNAs in nonrecombining Y chromosomes undergo lower rates of sequence evolution and homogenization than do satellite DNAs in autosomes.[Reviewing Editor: DR. Jerzy Jurka]     

Rapid Evolution by Positive Selection and Gene Gain and Loss: PLA<Subscript>2</Subscript> Venom Genes in Closely Related <Emphasis Type="Italic">Sistrurus</Emphasis> Rattlesnakes with Divergent Diets

Rapid evolution of snake venom genes by positive selection has been reported previously but key features of this process such as the targets of selection, rates of gene turnover, and functional diversity of toxins generated remain unclear. This is especially true for closely related species with divergent diets. We describe the evolution of PLA₂ gene sequences isolated from genomic DNA from four taxa of Sistrurus rattlesnakes which feed on different prey. We identified four to seven distinct PLA₂ sequences in each taxon and phylogenetic analyses suggest that these sequences represent a rapidly evolving gene family consisting of both paralogous and homologous loci with high rates of gene gain and loss. Strong positive selection was implicated as a driving force in the evolution of these protein coding sequences. Exons coding for amino acids that make up mature proteins have levels of variation two to three times greater than those of the surrounding noncoding intronic sequences. Maximum likelihood models of coding sequence evolution reveal that a high proportion (∼30%) of all codons in the mature protein fall into a class of codons with an estimated d _N /d _S (ω) ratio of at least 2.8. An analysis of selection on individual codons identified nine residues as being under strong (p < 0.01) positive selection, with a disproportionately high proportion of these residues found in two functional regions of the PLA₂ protein (surface residues and putative anticoagulant region). This is direct evidence that diversifying selection has led to high levels of functional diversity due to structural differences in proteins among these snakes. Overall, our results demonstrate that both gene gain and loss and protein sequence evolution via positive selection are important evolutionary forces driving adaptive divergence in venom proteins in closely related species of venomous snakes.     

Differential organellar inheritance in Passiflora’s (Passifloraceae) subgenera

Four chloroplast (cp), one mitochondrial (mt), and one ribosomal nuclear (ITS) DNA regions were studied in four artificial and one natural interspecific Passiflora hybrids. The ITS results confirmed their hybrid origin and all mtDNAs were maternally inherited. The same, however, was not true for cpDNA. The four hybrids (three artificial and one natural) derived from species of the Passiflora subgenus showed a cpDNA paternal inheritance, while the one involving taxa of the Decaloba subgenus gave evidence of maternal transmission. These results are of significance for the ongoing studies which are being performed on the molecular evolution of this genus and furnish important background for investigations aimed at clarifying the factors which determine cpDNA inheritance.     

Biochemical and immunochemical characterization of hop-hornbeam (Ostrya Carpinifolia Scop.) pollen

In the last few years Ostrya carpinifolia pollen is consideredas an important cause of respiratoryallergy in Mediterranean areas. The concentration ofthe pollen was measured over a period of fifteen yearsfrom 1981 to 1996 in an area around Genoa; the resultsof this study have clearly indicated an increasingtrend that correlate with persons sensitization.In this study we sought to define the immunochemical andbiochemical properties of hop-hornbean pollen. Soluble proteins extracted from Ostryacarpinifolia pollen and from the taxonomicallyrelated species Corylus Avellana, were analyzedby polyacrylamide gel electrophoresis (SDS-PAGE), byhorizontal isoelectrofocusing (IEF) and by twodimensions electrophoresis (2D-PAGE). Allergenicproteins were identified with sera of sensibilizedpatients and cross-reactivity was evaluated byimmunoblotting techniques. The electrophoreticanalysis showed a partial identity between theproteins from Ostrya and Corylus extracts. The immunoblotting assay, developed withhuman IgE from subjects allergic to hop-hornbeampollen, displayed the major IgE reactivity for acomponent with a molecular weight of 17 kDa expressedin both Ostrya and Corylus extracts. This reactivity is consistent with the presence ofBet v 1 that is described as the major pollen allergenin the Betulaceae and Corylaceae families. Sera fromsubjects allergic to Ostrya were then preadsorbed with recombinant Bet v 1 immobilized in the Pharmacia CAP System; a significant reduction ofthe IgE binding activity was observed after thetreatment. We therefore suggest that Bet v 1 couldbe one of the allergenic proteins present in theOstrya pollen possibly being responsible forcross-reactivity with other members of taxonomicallyrelated families.     

Concurrent evolution of nitrogenase genes and 16S rRNA in Rhizobium species and other nitrogen fixing bacteria

It was known that nitrogenase genes and proteins are well conserved even though they are present in a large variety of phylogenetically diverse nitrogen fixing bacteria. This has lead to the speculation, among others, that nitrogen fixation (nif) genes were spread by lateral gene transfer relatively late in evolution. Here we report an attempt to test this hypothesis.We had previously established the complete nucleotide sequences of the three nitrogenase genes from Bradyrhizobium japonicum, and have now analyzed their homologies (or the amino acid sequence homologies of their gene products) with corresponding genes (and proteins) from other nitrogen fixing bacteria. There was a considerable sequence conservation which certainly reflects the strict structural requirements of the nitrogenase iron-sulfur proteins for catalytic functioning. Despite this, the sequences were divergent enough to classify them into an evolutionary scheme that was conceptually not different from the phylogenetic positions, based on 16S rRNA homology, of the species or genera harboring these genes. Only the relation of nif genes of slow-growing rhizobia (to which B. japonicum belongs) and fast-growing rhizobia was unexpectedly distant. We have, therefore, performed oligonucleotide cataloguing of their 16S rRNA, and found that there was indeed only a similarity of S _AB=0.53 between fast- and slowgrowing rhizobia.In conclusion, the results suggest that nif genes may have evolved to a large degree in a similar fashion as the bacteria which carry them. This interpretation would speak against the idea of a recent lateral distribution of nif genes among microorganisms.     

Phylogenetic analysis of the thiolase family. Implications for the evolutionary origin of peroxisomes

Summary The thiolase family is a widespread group of proteins present in prokaryotes and three cellular compartments of eukaryotes. This fact makes this family interesting in order to study the evolutionary process of eukaryotes. Using the sequence of peroxisomal thiolase from Saccharomyces cerevisiae recently obtained by us and the other known thiolase sequences, a phylogenetic analysis has been carried out. It shows that all these proteins derived from a primitive enzyme, present in the common ancestor of eubacteria and eukaryotes, which evolved into different specialized thiolases confined to various cell compartments. The evolutionary tree obtained is compatible with the endosymbiotic theory for the origin of peroxisomes. Offprint requests to: J.E. Pérez-Ortín     

The major birch pollen allergen, Bet v 1, shows ribonuclease activity

The major birch (Betula alba L.) pollen allergen, Bet v 1, has been shown to be homologous to pathogenesis-related proteins in a number of plants. Recently, it was demonstrated that a ginseng protein with high homology to an intracellular pathogenesis-related protein of parsley and to Bet v 1 is a ribonuclease (RNase). Birch pollen extract was separated in an RNase activity gel. Four major RNase bands were excised from the gel, reseparated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and identified by Western blotting with a specific Bet v 1 monoclonal antibody and patient's serum. Thus the monomer and the dimer of Bet v 1 showed RNase activity. Purified recombinant Bet v 1 was shown to degrade plant RNA. The RNase activity of recombinant Bet v 1 was 180 units · mg^?1.     

The Evolutionary History of <Emphasis Type="Italic">Shigella</Emphasis> and Enteroinvasive <Emphasis Type="Italic">Escherichia coli</Emphasis> Revised

In Shigella and enteroinvasive Escherichia coli (EIEC), the etiologic agents of shigellosis in humans, the determinants responsible for entry of bacteria into and dissemination within epithelial cells are encoded by a virulence plasmid. To understand the evolution of the association between the virulence plasmid and the chromosome, we performed a phylogenetic analysis using the sequences of four chromosomal genes (trpA, trpB, pabB, and putP) and three virulence plasmid genes (ipaB, ipaD, and icsA) of a collection of 51 Shigella and EIEC strains. The phylogenetic tree derived from chromosomal genes showed a typical star phylogeny, indicating a fast diversification of Shigella and EIEC groups. Phylogenetic groups obtained from the chromosomal and plasmidic genes were similar, suggesting that the virulence plasmid and the chromosome share similar evolutionary histories. The few incongruences between the trees could be attributed to exchanges of fragments of different plasmids and not to the transfer of an entire plasmid. This indicates that the virulence plasmid was not transferred between the different Shigella and EIEC groups. These data support a model of evolution in which the acquisition of the virulence plasmid in an ancestral E. coli strain preceded the diversification by radiation of all Shigella and EIEC groups, which led to highly diversified but highly specialized pathogenic groups.     

Primate numts and reticulate evolution of capped and golden leaf monkeys (Primates: Colobinae)

A recent phylogenetic study of langurs and leaf monkeys of South Asia suggested a reticulate evolution of capped and golden leaf monkeys through ancient hybridization between Semnopithecus and Trachypithecus. To test this hybridization scenario, I analysed nuclear copies of the mitochondrial cytochrome b gene (numts) from capped, golden and Phayre’s leaf monkeys. These numts were aligned with mitochondrial cytochrome b sequences of various species belonging to the genera Semnopithecus and Trachypithecus. In the phylogenetic tree derived from this alignment, the numts fell into three distinct clades (A, B and C) suggesting three independent integration events. Clade A was basal to Semnopithecus, and clades B and C were basal to Trachypithecus. Among the numts in clades A and C were sequences derived from species not represented in their respective sister mitochondrial groups. This unusual placement of certain numts is taken as additional support for the hybridization scenario. Based on the molecular dating of these integration events, hybridization is estimated to have occurred around 7.1 to 3.4 million years ago. Capped and golden leaf monkeys might have to be assigned to a new genus to reconcile their unique evolutionary history. Additionally, northeast India appears to be a ‘hot spot’ for lineages that might have evolved through reticulate evolution.     

Characterization of PR-10 genes from eight Betula species and detection of Bet v 1 isoforms in birch pollen

Background  

Bet v 1 is an important cause of hay fever in northern Europe. Bet v 1 isoforms from the European white birch (Betula pendula) have been investigated extensively, but the allergenic potency of other birch species is unknown. The presence of Bet v 1 and closely related PR-10 genes in the genome was established by amplification and sequencing of alleles from eight birch species that represent the four subgenera within the genus Betula. Q-TOF LC-MS^E was applied to identify which PR-10/Bet v 1 genes are actually expressed in pollen and to determine the relative abundances of individual isoforms in the pollen proteome.     

Differential impact of retrotransposon populations on the genome of allotetraploid tobacco (<Emphasis Type="Italic">Nicotiana tabacum</Emphasis>)

LTR-retrotransposons contribute substantially to the structural diversity of plant genomes. Recent models of genome evolution suggest that retrotransposon amplification is offset by removal of retrotransposon sequences, leading to a turnover of retrotransposon populations. While bursts of amplification have been documented, it is not known whether removal of retrotransposon sequences occurs continuously, or is triggered by specific stimuli over short evolutionary periods. In this work, we have characterized the evolutionary dynamics of four populations of copia-type retrotransposons in allotetraploid tobacco (Nicotiana tabacum) and its two diploid progenitors Nicotiana sylvestris and Nicotiana tomentosiformis. We have used SSAP (Sequence-Specific Amplification Polymorphism) to evaluate the contribution retrotransposons have made to the diversity of tobacco and its diploid progenitor species, to quantify the contribution each diploid progenitor has made to tobacco's retrotransposon populations, and to estimate losses or amplifications of retrotransposon sequences subsequent to tobacco's formation. Our results show that the tobacco genome derives from a turnover of retrotransposon sequences with removals concomitant with new insertions. We have detected unique behaviour specific to each retrotransposon population, with differences likely reflecting distinct evolutionary histories and activities of particular elements. Our results indicate that the retrotransposon content of a given plant species is strongly influenced by the host evolutionary history, with periods of rapid turnover of retrotransposon sequences stimulated by allopolyploidy.     

Comparative analysis of avian BMAL1 and CLOCK protein sequences: a search for features associated with owl nocturnal behaviour

Animals differ widely in the phasing of their daily rhythms with respect to daily environmental rhythms. While birds are predominantly day-active, nocturnal activity is a characteristic feature of the order Strigiformes (owls). To study the evolution of owl night-activity cDNA sequences encoding the circadian core oscillator (CCO) proteins BMAL1 and CLOCK were obtained from barn owl (Tyto alba). The predicted proteins showed high sequence identity with their Galliform homologues (BMAL1: 99%; CLOCK: 95.6%). A computer-predicted chicken BMAL1 casein kinase-1 phosphorylation site is absent from T. alba BMAL1, but also absent from homologues of other six bird species (5 orders) (night-active (n=2), day-active (n=4)) indicating no evolutionary association with night activity. Sequence differences between T. alba and Galliform CLOCK frequently involved serine and threonine residues suggesting potential differences in their phosphorylation. The length of a poly-glutamine string in the CLOCK C-terminus varied between and within 25 species (6 orders) examined, however, no discernible feature distinguishing day and night active species was found. No differences were found between day (n=5) and night (n=7)-active species (12 species, 6 orders) in a region of the PER2 protein implicated in altered rhythm phasing in humans. In conclusion the avian CCO components examined showed strong evolutionary conservation. Molecular evolution associated with owl night-activity may have involved alterations in the CCO relationship with ‘output’ genes rather than in the molecular structure of the CCO itself.     

Genomic organization and evolution of the 5S ribosomal DNA in Tilapiini fishes

Summary 5S rDNA sequences present an intense dynamism and have proved to be valuable as genetic markers to distinguish closed related species and also in the understanding of the evolutionary dynamic of repetitive sequences in the genomes. In order to identify patterns of 5S rDNA organization and their evolution in the genome of fish species, such genomic segment was investigated in the tilapias Oreochromis niloticus and Tilapia rendalli, and in the hybrid O. urolepis hornorum × O. mossambicus. A dual 5S rDNA system was identified in the three analyzed tilapia samples. Although each 5S rDNA class was conserved among the three samples, a distinct 5S rDNA genome organization pattern could be evidenced for each sample. The presence of a dual 5S rDNA system seems to be a general trait among non-related teleost fish orders, suggesting that evolutionary events of duplication have occurred before the divergence of the main groups of teleost fishes.     

Veronica: Parallel morphological evolution and phylogeography in the Mediterranean

The genus Veronica s. lat. comprises about 450 species (including about 180 species from the southern hemisphere Hebe-complex), many of which grow in the Mediterranean area. Their extreme variability in morphology, life form and habitats has led to many suggestions regarding evolution and biogeography. Difficulties arise from parallel syndromes, widespread among alpine species and lowland perennials, and particularly among annual species of the genus. We have used sequences of the plastid trnL-F region and nuclear ribosomal ITS sequences to differentiate between different clades of Veronica and reveal cases of parallel evolution. Based on this data, cases of parallel evolution have been found in biogeographical patterns among the alpine species of Veroniceae, in which species from European mountains have affinities to those in the Central Asian/Himalayan region whereas alpine species from Turkey are probably more recently derived from lowland southwestern Asian taxa. Different subspecies of Veronica bombycina gained their characteristic morphology independently and parallel in adaptation to their alpine environment. Pinnatifid leaves have been gained parallel in perennial grassland species of Veronica. Finally, parallel evolutionary trends in many characters, not only morphological but also molecular characters, are common among annual species of Veronica.We wish to thank the Studienstiftung des deutschen Volkes for a doctoral scholarship to DCA and the Spanish Ministerio de Ciencia y Tecnologia for a postdoctoral grant (programa FPI) to MMO. This work was partially supported by the Junta de Castilla y Leon and the European Union (FSE) through the research project SA117/01. We also thank Prof. F. Ehrendorfer for critical comments and valuable discussions. Further, DCA thanks Daniela Hanfland and Yoshiki Nakamoto for help in the field. Manfred A. Fischer, Lena Struwe, Bernhard Dickore, Christoph Dobes, Eberhard Fischer, Niels Köster, Gerald Schneeweiss, Tod Stuessy, Tim Utteridge and Matsugo Yokota have provided plant material for this study.     

Phylogenetic and genetic studies in Papaver section Oxytona: cytogenetics,isozyme analysis and chloroplast DNA variation

Summary Chromosome behaviour at meiosis, isozyme studies and analysis of the chloroplast DNA restriction fragments were used to assess the phylogenetic relations among the three Papaver species of the section Oxytona. The multivalents observed in diplotene — diakinesis stages of meiosis of the hexaploid P. pseudo-orientale and its tetraploid hybrid with P. bracteatum indicate the autopolyploid nature of this section. Further evidence supporting this conclusion was obtained from isozyme analysis. The same number of isozymes was expressed in all the species regardless of their ploidy level. Inheritance studies conducted with Pgi, Dia and Acp allozymes demonstrated, for the first time, the transfer and expression of genetic material among these species. The differences found in the chloroplast DNA restriction fragments of the Oxytona species and of P. somniferum indicate intensive evolution of the chloroplast DNA in the genus Papaver. The similarity of the chloroplast DNA restriction patterns and of the isozymes in P. orientale and P. pseudo-orientale suggested that P. orientale was the female parent in the cross generating P. pseudo-orientale and that the latter species is of recent origin.Contribution no. 2199 —E, 1987 series from the Agricultural Research Organization, The Volcani center, Bet Dagan 50 250, Israel     

Speciation in <Emphasis Type="Italic">Chlamydia</Emphasis>: Genomewide Phylogenetic Analyses Identified a Reliable Set of Acquired Genes

Horizontal gene transfer (HGT), a process through which genomes acquire sequences from distantly related organisms, is believed to be a major source of genetic diversity in bacteria. A central question concerning the impact of HGT on bacterial genome evolution is the proportion of horizontally transferred sequences within genomes. This issue, however, remains unresolved because the various methods developed to detect potential HGT events identify different sets of genes. The present-day consensus is that phylogenetic analysis of individual genes is still the most objective and accurate approach for determining the occurrence and directionality of HGT. Here we present a genome-scale phylogenetic analysis of protein-encoding genes from five closely related Chlamydia, identifying a reliable set of sequences that have arisen via HGT since the divergence of the Chlamydia lineage. According to our knowledge, this is the first systematic phylogenetic inference-based attempt to establish a reliable set of acquired genes in a bacterial genome. Although Chlamydia are obligate intracellular parasites of higher eukaryotes, and thus suspected to be isolated from HGT more than the free-living species, our results show that their diversification has involved the introduction of foreign sequences into their genome. Furthermore, we also identified a complete set of genes that have undergone deletion, duplication, or rearrangement during this evolutionary period leading to the radiation of Chlamydia species. Our analysis may provide a deeper insight into how these medically important pathogens emerged and evolved from a common ancestor.     

The arms race between heliconiine butterflies and Passiflora plants – new insights on an ancient subject

Heliconiines are called passion vine butterflies because they feed exclusively on Passiflora plants during the larval stage. Many features of Passiflora and heliconiines indicate that they have radiated and speciated in association with each other, and therefore this model system was one of the first examples used to exemplify coevolution theory. Three major adaptations of Passiflora plants supported arguments in favour of their coevolution with heliconiines: unusual variation of leaf shape within the genus; the occurrence of yellow structures mimicking heliconiine eggs; and their extensive diversity of defence compounds called cyanogenic glucosides. However, the protection systems of Passiflora plants go beyond these three features. Trichomes, mimicry of pathogen infection through variegation, and production of extrafloral nectar to attract ants and other predators of their herbivores, are morphological defences reported in this plant genus. Moreover, Passiflora plants are well protected chemically, not only by cyanogenic glucosides, but also by other compounds such as alkaloids, flavonoids, saponins, tannins and phenolics. Heliconiines can synthesize cyanogenic glucosides themselves, and their ability to handle these compounds was probably one of the most crucial adaptations that allowed the ancestor of these butterflies to feed on Passiflora plants. Indeed, it has been shown that Heliconius larvae can sequester cyanogenic glucosides and alkaloids from their host plants and utilize them for their own benefit. Recently, it was discovered that Heliconius adults have highly accurate visual and chemosensory systems, and the expansion of brain structures that can process such information allows them to memorize shapes and display elaborate pre‐oviposition behaviour in order to defeat visual barriers evolved by Passiflora species. Even though the heliconiine–Passiflora model system has been intensively studied, the forces driving host‐plant preference in these butterflies remain unclear. New studies have shown that host‐plant preference seems to be genetically controlled, but in many species there is some plasticity in this choice and preferences can even be induced. Although much knowledge regarding the coevolution of Passiflora plants and heliconiine butterflies has accumulated in recent decades, there remain many exciting unanswered questions concerning this model system.